Contamination of blood supplies with infectious microorganisms such as HIV, hepatitis and other viruses and bacteria presents a serious health hazard for those who must receive transfusions of whole blood or administration of various blood components such as platelets, red cells, blood plasma, Factor VIII, plasminogen, fibronectin, anti-thrombin III, cryoprecipitate, human plasma protein fraction, albumin, immune serum globulin, prothrombin complex plasma growth hormones, and other components isolated from blood. Blood screening procedures currently available may miss contaminants. Thus, there is a need for sterilization procedures that effectively neutralize all infectious viruses and other microorganisms but do not damage cellular blood components, do not degrade desired biological activities of proteins, and preferably do not need to be removed prior to administration of the blood product to the patient.
The use of photosensitizers, compounds which absorb light of a defined wavelength and transfer the absorbed energy to an energy acceptor, has been proposed for blood component sterilization. Various photosensitizers have been proposed for use as blood additives. A review of some photosensitizers including psoralens, and some of the issues of importance in choosing photosensitizers for decontamination of blood products is provided in Goodrich, R. P., et al. (1997), xe2x80x9cThe Design and Development of Selective, Photoactivated Drugs for Sterilization of Blood Products,xe2x80x9d Drugs of the Future 22:159-171.
Some photosensitizers that have been proposed for use for blood component sterilization have undesirable properties. For example, European Patent Application 196,515 published Oct. 8, 1986, suggests the use of non-endogenous photosensitizers such as porphyrins, psoralens, acridine, toluidines, flavine (acriflavine hydrochloride), phenothiazine derivatives, and dyes such as neutral red and methylene blue, as blood additives. Another molecule, chlorpromazine, has been used as a photosensitizer; however its usefulness is limited by the fact that it should be removed from any fluid administered to a patient after the decontamination procedure because it has a sedative effect. Protoporphyrin, which occurs naturally within the body, can be metabolized to form a photosensitizer; however, its usefulness is limited in that it degrades the desired biological activities of proteins.
In addition to molecules which can serve as photosensitizers, alkylating agents have been proposed for use as blood contaminant neutralizers. Aklylating agents are believed to deactivate microorganisms by alkylating nucleophilic groups of amino acid residues and nucleic bases at a certain pH. Ethyleneimine has been reported to deactivate certain viruses (U.S. Pat. No. 5,891,075 (Budowsky, et al.), WO 97/107674 (published Mar. 6, 1997)).
U.S. patent application Ser. No. 09/119,666 and continuation in part Ser. No. 09/357,188, hereby incorporated by reference to the extent not inconsistent with the disclosure herein, describes methods and apparatus for neutralization of biological contaminants using endogenous photosensitizers, including 7,8-dimethyl-10-ribityl isoalloxazine (riboflavine). 
7,8-dimethyl-10-ribityl isoalloxazine (Riboflavine or vitamin B2) absorbs light from about 200 to 500 nm. The ring system core of 7,8-dimethyl-10-ribityl isoalloxazine is resistant to photodegradation but the ribityl side chain of riboflavin undergoes photodegradation. Photolysis of 7,8-dimethyl-10-ribityl isoalloxazine may form lumichrome (7,8-dimethylalloxazine) depending on conditions. 7,8-dimethylalloxazine strongly absorbs ultraviolet (UV) light and only weakly absorbs visible light. 
U.S. Pat. No. 5,811,144 discusses the treatment of beer with visible light under substantially anaerobic conditions to reportedly reduce the riboflavin content of the beer.
Small molecules such as those shown below which are derived from the ribityl side chain are expected to be products from the photolysis of riboflavin. 
Incomplete photolysis of riboflavin leads to isoalloxazine-containing intermediates (Smith, E. C. and Metzler, D. E. (1963) J. Am. Chem. Soc. 85:3285-3288; Carins, W. L. and Metzler, D. E. (1971) J. Am. Chem. Soc. 93:2772-2777; Treadwell, G. E. et al. (1968) J. Chromatog. 35:376-388). Some of the identified compounds are: 
These compounds absorb visible light and may convert to either lumichrome or another riboflavin metabolite, lumiflavin (7,8,10-trimethylisoalloxazine) upon complete photolysis, depending on the experimental conditions. 
Lumichrome and lumiflavin are reported to be produced by the photolysis of milk (Parks, O. W. and Allen, C. (1977) Dairy Sci. 61:1038-1041; Toyosaki, T. and Hayashi, A. (1993) Milewissenschaft 48:607-609).
As a result of the degradation of 7,8-dimethyl-10-ribityl isoalloxazine upon exposure to light, a combination of visible and ultraviolet light is preferred in decontamination procedures using 7,8-dimethyl-10-ribityl isoalloxazine. Since UV light has a higher energy per photon than visible light, and because UV light is absorbed more strongly than visible light by useful compounds in the biological fluid, more damage to the useful components in the biological fluid containing the contaminants will occur when ultraviolet light is used in combination with visible light than when visible light can be used alone.
There is a need for compounds that neutralize microorganisms with visible light alone.
All publications referred to herein are hereby incorporated by reference to the extent not inconsistent with the disclosure herein.
Methods are provided for treating a fluid or other material to neutralize at least some of the microorganisms and white cells which may be present therein or thereon. Such fluids may also contain one or more components selected from the group consisting of protein, e.g. biologically active protein such as a therapeutic protein, blood and blood constituents, without destroying the biological activity of such components. The methods comprise:
(a) mixing a neutralization-effective amount of a microorganism neutralizer of formula: 
with the fluid, wherein R1, R2, R3, R4, R5 and R6 are, independently from one another, selected from the group consisting of hydrogen, optionally substituted hydrocarbyl, alcohol, amine, polyamine, sulfate, phosphate, halogen selected from the group consisting of chlorine, bromine and iodine, salts of the foregoing, and xe2x80x94NRaxe2x80x94(CRbRc)nxe2x80x94X wherein X is a halogen selected from the group consisting of chlorine, bromine and iodine, Ra, Rb and Rc are, independently of each other, selected from the group consisting of hydrogen, optionally substituted hydrocarbyl, and halogen selected from the group consisting of chlorine, bromine and iodine, and n is an integer from 0 to 20;
provided that R1 is not xe2x80x94OH or a straight chain alkyl group where the second carbon of the chain is substituted with xe2x80x94OH or xe2x95x90O except that the compound may be 
and provided that R1, R4 and R5 are not all methyl groups when R2, R3 and R6 are all hydrogen;
(b) exposing the fluid to a triggering event, whereby at least some of the microorganisms are neutralized.
In one group of compounds, n is an integer between 0 and 5. In another group of compounds, n is an integer from 0 to 10. In another group of compounds, n is an integer from 0 to 20.
One group of compounds include the following: 
where R5 is specifically shown as methyl, but may be H or C1-C3 alkyl chains; and 
where R4 is specifically shown as methyl, but may be H or C1-C3 alkyl chains.
A fluid is provided comprising biologically active protein, blood or blood constituents, and microorganism neutralizer, made by the method above. The fluid may also contain neutralized microorganisms. A blood product is also provided comprising a microorganism neutralizer made by the method above.
Compounds are provided having the structure: 
wherein R1, R2, R3, R4, R5 and R6 are, independently from one another, selected from the group consisting of hydrogen, optionally substituted hydrocarbyl, alcohol, amine, polyamine, sulfate, phosphate, halogen selected from the group consisting of chlorine, bromine and iodine, salts of the foregoing, and xe2x80x94NRaxe2x80x94(CRbRc)nxe2x80x94X wherein X is a halogen selected from the group consisting of chlorine, bromine and iodine, Ra, Rb and Rc are, independently of each other, selected from the group consisting of hydrogen, optionally substituted hydrocarbyl, and halogen selected from the group consisting of chlorine, bromine and iodine, and n is an integer from 0 to 20;
provided that R1is not xe2x80x94OH or a straight chain alkyl group where the second carbon of the chain is substituted with xe2x80x94OH or xe2x95x90O; except that the compound may be 
and provided that R1 is not a 2-, 3-, 4- or 5-carbon straight chain alkyl that terminates in xe2x80x94OH, xe2x80x94COH, or xe2x80x94H when R2, R3 and R6 are H, and R4 and R5 are CH3; R1 is not xe2x80x94CH2CH2xe2x80x94(CHOH)2xe2x80x94CH3 or xe2x80x94CH2CH2xe2x80x94(CHOH)2xe2x80x94CH2SO4 or 1xe2x80x2-D-sorbityl or 1xe2x80x2-D-dulcityl or 1xe2x80x2-D-rhamnityl or 1xe2x80x2-D,L-glyceryl or xe2x80x94CH2xe2x80x94Oxe2x80x94C(O)xe2x80x94CH3 or xe2x80x94CH2xe2x80x94Oxe2x80x94C(O)xe2x80x94CH2CH3 or 2xe2x80x2,3xe2x80x2,4xe2x80x2,5xe2x80x2-di-O-isopropyridene-riboflavin or 8-aminooctyl when R2, R3 and R6 are H and R4 and R5 are CH3; R1 is not 1xe2x80x2-D-sorbityl or 1xe2x80x2-D-dulcityl when R4 and R5 are both chlorines and when R2, R3 and R6 are all hydrogens; R5 is not ethyl or chloro when R1 and R4 are methyl and R2, R3 and R6 are all hydrogens; R4 and R5 are not both methoxy or both tetramethylene when R1 is methyl and R2, R3 and R6 are all hydrogens; R2 is not xe2x80x94CH2CH2NH when R1, R4 and R5 are CH3 and R3 and R6 are H; R2 is not 
when R1, R4 and R5 are CH3 and R3 and R6 are H; R5 is not chloro when R4 is methoxy and R1 is ethyl-2xe2x80x2N-pyrrolidino and R2, R3, and R6 are hydrogen; R1 is not N,N-dimethylaminopropyl or N,N-diethylaminoethyl when R5 is chloro or methyl and R2, R3, R4 and R6 are hydrogen; R3 is not xe2x80x94NH(CH2CH2)Cl when R6 is xe2x80x94NH2 and R1, R2, R4 and R5 are H; R1, R4, R5 are not all methyl groups when all of R2, R3 and R6 are hydrogens; R1, R4, R5 and R2 are not all methyl groups when R3 and R6 are hydrogens; R2 is not carboxymethyl when R1, R4 and R5 are methyl and R3 and R6 are hydrogen; R4 is not xe2x80x94NH2 when R1 and R5 are methyl and R2, R3 and R6 are all hydrogen; R1 is not a phenyl group when R4 and R5 are methyl and R2, R3 and R6 are all H; R1 is not methyl or N,N-dimethylaminoethyl when all of R2, R3, R4, R5 and R6 are hydrogen; R2, R4, R5 are not all methyl when R1 is acetoxyethyl and R3 and R6 are hydrogen; R5 is not methyl when R1 is N,N-diethylaminoethyl and R2, R3, R4 and R6 are all hydrogen; R4 and R5 are not both chlorine when R1 is methyl and R2, R3 and R6 are all hydrogen; R1 is not ethyl, xcex2-chloroethyl, n-butyl, anilino, benzyl, phenyl, p-tolyl or p-anisyl when R5 is NH2 and R2, R3, R4 and R6 are all hydrogen; and R4 is not chlorine when R1 is N,N-dimethylaminopropyl and R2, R3, R5 and R6 are all hydrogen.
In one group of compounds, n is an integer between 0 and 5. In another group of compounds, n is an integer from 0 to 10. In another group of compounds, n is an integer from 0 to 20.
Compounds containing any combination of substituents or members of the Markush groups specified above are within the scope of the invention. All compounds of the invention have the ability to neutralize microorganisms. All substituents of the compounds of the invention may be the same, all substituents may be different, or any combination of substituents may be the same or different. Substituents with a specified function, for example those that impart water solubility to the compound, may be included at any of R1-R6. Compounds of the invention include all those compounds with the isoalloxazine backbone (shown below): 
where R1-R6 are substituted with various substituents, as described elsewhere, except those previously known to the art. The substituents included in the compounds and used in the methods of the invention may be any substituent not having structures or reactivity which would substantially interfere with the desired microorganism neutralization of the microorganism neutralizer, as may readily be determined without undue experimentation by those skilled in the art.
The invention provides a class of compounds wherein a plurality of R1, R2, R3, R4, R5 and R6 are neither CH3 nor H; and a class of compounds wherein one of R1, R2, R3, R4, R5 and R6 is neither CH3 nor H. Particular embodiments of compounds of those classes include those wherein a R1, R2, R3, R4, R5 or R6 which is neither CH3 nor H imparts substantial water solubility to the microorganism neutralizer. Preferred examples of these compounds are: 
wherein R is a substituent imparting water solubility to the molecule, including, but not limited to, ascorbate, alcohol, polyalcohol; amine or polyamines, straight chain or cyclic saccharides, sulfates, phosphates, alkyl chains optionally substituted with xe2x80x94OH at any position, glycols, including polyethylene glycol and polyethers.
Another class of compounds of the invention include those wherein a R1, R2, R3, R4, R5 or R6 that is neither H nor CH3 contains a halogen or is a halogen, wherein the halogen is selected from the group consisting of fluorine, chlorine, bromine and iodine. Particular embodiments of compounds of this class include compounds where a R1, R2, R3, R4, R5 or R6 that is neither H nor CH3 is: xe2x80x94NRaxe2x80x94(CRbRc)nxe2x80x94X wherein X is a halogen selected from the group consisting of chlorine, bromine and iodine, or is a water soluble group, Ra, Rb and Rc are, independently of each other, selected from the group consisting of hydrogen and optionally substituted hydrocarbyl, and n is an integer from 0 to 20.
Preferred examples of compounds of this class are: 
where W is a substituent imparting water solubility to the molecule, including, but not limited to, ascorbate, alcohol, polyalcohol; amine or polyamines, straight chain or cyclic saccharides, sulfates, phosphates, alkyl chains optionally substituted with xe2x80x94OH at any position, glycols, including polyethylene glycol and polyethers.
Another particular embodiment of compounds wherein a R1, R2, R3, R4, R5 or R6 that is neither H nor CH3 contains a halogen or is a halogen includes compounds wherein a R1, R2, R3, R4, R5 or R6 that is neither H nor CH3 is: Xxe2x80x94(CH2)nxe2x80x94, wherein X is a halogen selected from the group consisting of chlorine, bromine and iodine, and n is an integer from 0 to 6. A preferred example of compounds of this class include: 
Other classes of compounds of this invention include those wherein R1 is CH2xe2x80x94(CH2OH)3xe2x80x94CH2OH and those wherein R1 is not CH2xe2x80x94(CH2OH)3xe2x80x94CH2OH. Also, those compounds wherein R3 and R6 are H are included in the invention.
A xe2x80x9ccarbonyl compoundxe2x80x9d is any compound containing a carbonyl group (xe2x80x94Cxe2x95x90O). The term xe2x80x9caminexe2x80x9d refers to a primary, secondary, or tertiary amine group. A xe2x80x9cpolyaminexe2x80x9d is a group that contains more than one amine group. A xe2x80x9csulfatexe2x80x9d group is a salt of sulfuric acid. Sulfate groups include the group (SO4)2xe2x88x92. xe2x80x9cPhosphatesxe2x80x9d contain the group PO43xe2x88x92. xe2x80x9cGlycolsxe2x80x9d are groups that have two alcohol groups per molecule of the compound. xe2x80x9cGlycolsxe2x80x9d are also known as diols. A glycol is described by the formula: CnH2n(OH)2, where n is an integer. An xe2x80x9caldehydexe2x80x9d is a group containing the formula xe2x80x94(Cxe2x95x90O)xe2x80x94H. A xe2x80x9cketonexe2x80x9d is a group with formula Rxe2x80x94(Cxe2x95x90O)xe2x80x94R, where R is not hydrogen. The R groups on ketones do not need to be the same. A xe2x80x9ccarboxylic acidxe2x80x9d is a group which includes the formula: xe2x80x94COOH. An xe2x80x9cetherxe2x80x9d is a group containing xe2x80x94Oxe2x80x94. A xe2x80x9csaltxe2x80x9d is a group where a hydrogen atom of an acid has been replaced with a metal atom or a positive radical, such as NH4+. xe2x80x9cAscorbatexe2x80x9d includes groups with formula: 
The term xe2x80x9chydrocarbylxe2x80x9d is used herein to refer generally to organic groups comprised of carbon chains to which hydrogen and optionally other elements are attached. CH2 or CH groups and C atoms of the carbon chains of the hydrocarbyl may be replaced with one or more heteroatoms (i.e., non-carbon atoms). Suitable heteroatoms include but are not limited to O, S, P and N atoms. The term hydrocarbyl includes, but is not limited to alkyl, alkenyl, alkynyl, ether, polyether, thioether, straight chain or cyclic saccharides, ascorbate, aminoalkyl, hydroxylalkyl, thioalkyl, aryl and heterocyclic aryl groups, optionally substituted isoalloxazine molecules, amino acid, polyalcohol, glycol, groups which have a mixture of saturated and unsaturated bonds, carbocyclic rings and combinations of such groups. The term also includes straight-chain, branched-chain and cyclic structures or combinations thereof. Hydrocarbyl groups are optionally substituted. Hydrocarbyl substitution includes substitution at one or more carbons in the group by moieties containing heteroatoms. Suitable substituents for hydrocarbyl groups include but are not limited to halogens, including chlorine, fluorine, bromine and iodine, OH, SH, NH2, COH, CO2H, ORa, SRa, NRaRb, CONRaRb, where Ra and Rb independently are alkyl, unsaturated alkyl or aryl groups.
The term xe2x80x9calkylxe2x80x9d takes its usual meaning in the art and is intended to include straight-chain, branched and cycloalkyl groups. The term includes, but is not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, neopentyl, 2-methylbutyl, 1-methylbutyl, 1-ethylpropyl, 1,1-dimethylpropyl, n-hexyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1-dimethylbutyl, 2-ethylbutyl, 1-ethylbutyl, 1,3-dimethylbutyl, n-heptyl, 5-methylhexyl, 4-methylhexyl, 3-methylhexyl, 2-methylhexyl, 1-methylhexyl, 3-ethylpentyl, 2-ethylpentyl, 1-ethylpentyl, 4,4-dimethylpentyl, 3,3-dimethylpentyl, 2,2-dimethylpentyl, 1,1-dimethylpentyl, n-octyl, 6-methylheptyl, 5-methylheptyl, 4-methylheptyl, 3-methylheptyl, 2-methylheptyl, 1-methylheptyl, 1-ethyhexyl, 1-propylpentyl, 3-ethylhexyl, 5,5-dimethylhexyl, 4,4-dimethylhexyl, 2,2-diethylbutyl, 3,3-diethylbutyl, and 1-methyl-1-propylbutyl. Alkyl groups are optionally substituted. Lower alkyl groups are C1-C6 alkyl and include among others methyl, ethyl, n-propyl, and isopropyl groups.
The term xe2x80x9ccycloalkylxe2x80x9d refers to alkyl groups having a hydrocarbon ring, particularly to those having rings of 3 to 7 carbon atoms. Cycloalkyl groups include those with alkyl group substitution on the ring. Cycloalkyl groups can include straight-chain and branched-chain portions. Cycloalkyl groups include but are not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and cyclononyl. Cycloalkyl groups can optionally be substituted.
Aryl groups may be substituted with one, two or more simple substituents including, but not limited to, lower alkyl, e.g., methyl, ethyl, butyl; halo, e.g., chloro, bromo; nitro; sulfato; sulfonyloxy; carboxy; carbo-lower-alkoxy, e.g., carbomethoxy, carbethoxy; amino; mono- and di-lower-alkylamino, e.g., methylamino, ethylamino, dimethylamino, methylethylamino; amido; hydroxy; lower-alkoxy, e.g., methoxy, ethoxy; and lower-alkanoyloxy, e.g., acetoxy.
The term xe2x80x9cunsaturated alkylxe2x80x9d group is used herein generally to include alkyl groups in which one or more carbon-carbon single bonds have been converted to carbon-carbon double or triple bonds. The term includes alkenyl and alkynyl groups in their most general sense. The term is intended to include groups having more than one double or triple bond, or combinations of double and triple bonds. Unsaturated alkyl groups include, without limitation, unsaturated straight-chain, branched or cycloalkyl groups. Unsaturated alkyl groups include without limitation: vinyl, allyl, propenyl, isopropenyl, butenyl, pentenyl, hexenyl, hexadienyl, heptenyl, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadienyl, cyclohexenyl, cyclohexadienyl, 1-propenyl, 2-butenyl, 2-methyl-2-butenyl, ethynyl, propargyl, 3-methyl-1-pentynyl, and 2-heptynyl. Unsaturated alkyl groups can optionally be substituted.
Substitution of alkyl, cycloalkyl and unsaturated alkyl groups includes substitution at one or more carbons in the group by moieties containing heteroatoms. Suitable substituents for these groups include but are not limited to OH, SH, NH2, COH, CO2H, ORc, SRc, P, PO, NRcRd, CONRcRd, and halogens, particularly chlorines and bromines where Rc and Rd, independently, are alkyl, unsaturated alkyl or aryl groups. Preferred alkyl and unsaturated alkyl groups are the lower alkyl, alkenyl or alkynyl groups having from 1 to about 3 carbon atoms.
The term xe2x80x9carylxe2x80x9d is used herein generally to refer to aromatic groups which have at least one ring having a conjugated pi electron system and includes without limitation carbocyclic aryl, aralkyl, heterocyclic aryl, biaryl groups and heterocyclic biaryl, all of which can be optionally substituted. Preferred aryl groups have one or two aromatic rings.
xe2x80x9cCarbocyclic arylxe2x80x9d refers to aryl groups in which the aromatic ring atoms are all carbons and includes without limitation phenyl, biphenyl and napthalene groups.
xe2x80x9cAralkylxe2x80x9d refers to an alkyl group substituted with an aryl group. Suitable aralkyl groups include among others benzyl, phenethyl and picolyl, and may be optionally substituted. Aralkyl groups include those with heterocyclic and carbocyclic aromatic moieties.
xe2x80x9cHeterocyclic aryl groupsxe2x80x9d refers to groups having at least one heterocyclic aromatic ring with from 1 to 3 heteroatoms in the ring, the remainder being carbon atoms. Suitable heteroatoms include without limitation oxygen, sulfur, and nitrogen. Heterocyclic aryl groups include among others furanyl, thienyl, pyridyl, pyrrolyl, N-alkyl pyrrolo, pyrimidyl, pyrazinyl, imidazolyl, benzofuranyl, quinolinyl, and indolyl, all optionally substituted.
xe2x80x9cHeterocyclic biarylxe2x80x9d refers to heterocyclic aryls in which a phenyl group is substituted by a heterocyclic aryl group ortho, meta or para to the point of attachment of the phenyl ring to the decalin or cyclohexane. Heterocyclic biaryl includes among others groups which have a phenyl group substituted with a heterocyclic aromatic ring. The aromatic rings in the heterocyclic biaryl group can be optionally substituted.
xe2x80x9cBiarylxe2x80x9d refers to carbocyclic aryl groups in which a phenyl group is substituted by a carbocyclic aryl group ortho, meta or para to the point of attachment of the phenyl ring to the decalin or cyclohexane. Biaryl groups include among others a first phenyl group substituted with a second phenyl ring ortho, meta or para to the point of attachment of the first phenyl ring to the decalin or cyclohexane structure. Para substitution is preferred. The aromatic rings in the biaryl group can be optionally substituted.
Aryl group substitution includes substitutions by non-aryl groups (excluding H) at one or more carbons or where possible at one or more heteroatoms in aromatic rings in the aryl group. Unsubstituted aryl, in contrast, refers to aryl groups in which the aromatic ring carbons are all substituted with H, e.g. unsubstituted phenyl (xe2x80x94C6H5), or naphthyl (xe2x80x94C10H7). Suitable substituents for aryl groups include among others, alkyl groups, unsaturated alkyl groups, halogens, OH, SH, NH2, COH, CO2H, ORc, SRc, NRcRf, CONRcRf, where Rc and Rf independently are alkyl, unsaturated alkyl or aryl groups. Preferred substituents are OH, SH, ORc, and SRc, where Rc is a lower alkyl, i.e., an alkyl group having from 1 to about 3 carbon atoms. Other preferred substituents are halogens, more preferably chlorine or bromine, and lower alkyl and unsaturated lower alkyl groups having from 1 to about 3 carbon atoms. Substituents include bridging groups between aromatic rings in the aryl group, such as xe2x80x94CO2xe2x80x94, xe2x80x94COxe2x80x94, xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94Pxe2x80x94, xe2x80x94NHxe2x80x94, xe2x80x94CHxe2x95x90CHxe2x80x94 and xe2x80x94(CH2)lxe2x80x94 where l is an integer from 1 to about 5, and particularly xe2x80x94CH2xe2x80x94. Examples of aryl groups having bridging substituents include phenylbenzoate. Substituents also include moieties, such as xe2x80x94(CH2)lxe2x80x94, xe2x80x94Oxe2x80x94(CH2)lxe2x80x94 or xe2x80x94OCOxe2x80x94(CH2)lxe2x80x94, where l is an integer from about 2 to 7, as appropriate for the moiety, which bridge two ring atoms in a single aromatic ring as, for example, in a 1,2,3,4-tetrahydronaphthalene group. Alkyl and unsaturated alkyl substituents of aryl groups can in turn optionally be substituted as described supra for substituted alkyl and unsaturated alkyl groups.
The terms xe2x80x9calkoxy groupxe2x80x9d and xe2x80x9cthioalkoxy groupxe2x80x9d (also known as mercaptide groups, the sulfur analog of alkoxy groups) take their generally accepted meaning. Alkoxy groups include but are not limited to methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, isobutoxy, tert-butoxy, n-pentyloxy, neopentyloxy, 2-methylbutoxy, 1-methylbutoxy, 1-ethyl propoxy, 1,1-dimethylpropoxy, n-hexyloxy, 1-methylpentyloxy, 2-methylpentyloxy, 3-methylpentyloxy, 4-methylpentyloxy, 3,3-dimethylbutoxy, 2,2-dimethoxybutoxy, 1-1-dimethylbutoxy, 2-ethylbutoxy, 1-ethylbutoxy, 1,3-dimethylbutoxy, n-pentyloxy, 5-methylhexyloxy, 4-methylhexyloxy, 3-methylhexyloxy, 2-methylhexyloxy, 1-methylhexyloxy, 3-ethylpentyloxy, 2-ethylpentyloxy, 1-ethylpentyloxy, 4,4-dimethylpentyloxy, 3,3-dimethylpentyloxy, 2,2-dimethylpentyloxy, 1,1-dimethylpentyloxy, n-octyloxy, 6-methylheptyloxy, 5-methylheptyloxy, 4-methylheptyloxy, 3-methylheptyloxy, 2-methylheptyloxy, 1-methylheptyloxy, 1-ethylhexyloxy, 1-propylpentyloxy, 3-ethylhexyloxy, 5,5-dimethylhexyloxy, 4,4-dimethylhexyloxy, 2,2-diethylbutoxy, 3,3-diethylbutoxy, 1-methyl-1-propylbutoxy, ethoxymethyl, n-propoxymethyl, isopropoxymethyl, sec-butoxymethyl, isobutoxymethyl, (1-ethyl propoxy)methyl, (2-ethylbutoxy)methyl, (1-ethylbutoxy)methyl, (2-ethylpentyloxy)methyl, (3-ethylpentyloxy)methyl, 2-methoxyethyl, 1-methoxyethyl, 2-ethoxyethyl, 3-methoxypropyl, 2-methoxypropyl, 1-methoxypropyl, 2-ethoxypropyl, 3-(n-propoxy)propyl, 4-methoxybutyl, 2-methoxybutyl, 4-ethoxybutyl, 2-ethoxybutyl, 5-ethoxypentyl, and 6-ethoxyhexyl. Thioakoxy groups include but are not limited to the sulfur analogs of the alkoxy groups specifically listed supra.
xe2x80x9cOptionalxe2x80x9d or xe2x80x9coptionallyxe2x80x9d means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances in which it does not. For example, xe2x80x9coptionally substituted phenylxe2x80x9d means that the phenyl radical may or may not be substituted and that the description includes both unsubstituted phenyl radicals and phenyl radicals wherein there is substitution.
xe2x80x9cAmino acidsxe2x80x9d as used herein include naturally occurring and commercially available amino acids and optical isomers thereof. Typical natural and commercially available amino acids are glycine, alanine, serine, homoserine, threonine, valine, norvaline, leucine, isoleucine, norleucine, aspartic acid, glutamic acid, lysine, ornithine, histidine, arginine, cysteine, homocysteine, methionine, phenylalanine, homophenylalanine, phenylglycine, o-, m-, and p-tyrosine, tryptophan, glutamine, asparagine, proline and hydroxyproline. xe2x80x9cAmino acidxe2x80x9d as used herein includes amino acid residues and amino acid side chains. An xe2x80x9camino acid residuexe2x80x9d is an amino acid radical xe2x80x94NHCH(R)C(O)xe2x80x94, wherein R is an amino acid side chain, except for the amino acid residues of proline and hydroxyproline which are xe2x80x94N(CH2xe2x80x94CH2xe2x80x94CH2)CHC(O)xe2x80x94 and xe2x80x94N(CHxe2x80x94CHOHCH2)CHC(O)xe2x80x94, respectively. An amino acid side chain is a radical found on the xcex1-carbon of an xcex1-amino acid as defined herein, where the radical is either hydrogen (side chain of glycine), methyl (side chain of alanine), or is a radical bonded to the xcex1-carbon by a methylene (xe2x80x94CH2xe2x80x94), or phenyl group.
A protected glucose derivative takes its usual meaning in the art and includes a glucose molecule wherein some of the hydroxyl groups are substituted with acetate groups.
xe2x80x9cContactingxe2x80x9d reaction components with each other refers to providing a medium and/or reaction chamber in which the reaction components are placed together so that they can react with each other. Preferably, the reaction components are suspended or dissolved in a carrier fluid which is a liquid medium. xe2x80x9cMaintaining reaction components in contactxe2x80x9d means keeping the components together in such a way that they can react with each other.
xe2x80x9cStraight chain or cyclic saccharidesxe2x80x9d include mono-, di- and poly-, straight chain and cyclic saccharides that are optionally substituted with an amino group which is optionally acetylated. Straight chain saccharides that are useful in this invention include but are not limited to those molecules with a chain of 5 or 6 carbon atoms with one or more xe2x80x94OH groups attached, and either an aldehyde or ketone group. Cyclic saccharides are saccharides that are in a ring form. Disaccharides are compounds wherein two monosaccharide groups are linked. Polysaccharides are compounds wherein more than two monosaccharide groups are linked. Specific examples of saccharides useful in this invention include glucose, ribose and glucosamine, among others.
xe2x80x9cIsoalloxazinexe2x80x9d, xe2x80x9cisoalloxazine derivativexe2x80x9d or xe2x80x9ccore structure of isoalloxazinexe2x80x9d include compounds that comprise the structure: 
where R1-R6 are substituted with various substituents, as described elsewhere.
As used herein, the term xe2x80x9cneutralization of a microorganismxe2x80x9d or xe2x80x9cneutralizingxe2x80x9d means totally or partially preventing the microorganism from replicating, either by killing the microorganism or otherwise interfering with its ability to reproduce. A xe2x80x9cneutralizerxe2x80x9d is a compound that is capable of neutralizing a microorganism. The neutralizers useful in this invention include molecules with the core structure of isoalloxazine, as defined above. To xe2x80x9cactivate the microorganism neutralizerxe2x80x9d is to expose the microorganism neutralizer to a triggering event that causes it to become active toward neutralizing microorganisms.
Microorganisms include viruses (both extracellular and intracellular), bacteria, bacteriophages, fungi, blood-transmitted parasites, and protozoa Exemplary viruses include acquired immunodeficiency (HIV) virus, hepatitis A, B and C viruses, sinbis virus, cytomegalovirus, vesicular stomatitis virus, herpes simplex viruses, e.g. types I and II, human T-lymphotropic retroviruses, HTLV-III, lymphadenopathy virus LAV/IDAV, parvovirus, transfusion-transmitted (TT) virus, Epstein-Barr virus, and others known to the art. Bacteriophages include "PHgr"X174, "PHgr"6, xcex, R17, T4, and T2. Exemplary bacteria include P. aeruginosa, S. aureus, S. epidermis, L. monocytogenes, E. coli, K. pneumonia and S. marcescens. Neutralization of white blood cells may be desirable when suppression of immune or autoimmune response is desired, e.g., in processes involving transfusion of red cells, platelets or plasma when donor white blood cells may be present.
xe2x80x9cTriggering eventxe2x80x9d refers to the stimulus that activates the microorganism neutralizer. Preferred triggering events include exposure of the neutralizer to an neutralization effective wavelength of light, or a pH sufficient to activate the neutralizer to neutralize microorganisms.
xe2x80x9cWater soluble groupxe2x80x9d includes a group that, when included as a substituent on the neutralizer, imparts substantial solubility in water to the compound. Typically, the compound is soluble in water at a concentration of about 10-150xcexcM. Water soluble groups as referred to in this invention include, but are not limited to alcohols; polyalcohols, straight chain or cyclic saccharides; amines and polyamines; sulfate groups; phosphate groups; ascorbate groups; alkyl chains optionally substituted with xe2x80x94OH at any position; glycols, including polyethylene glycols, and polyethers.
The term xe2x80x9cbiologically activexe2x80x9d means capable of effecting a change in a living organism or component thereof. xe2x80x9cBiologically activexe2x80x9d with respect to xe2x80x9cbiologically active proteinxe2x80x9d as referred to herein does not refer to proteins which are part of the microorganisms being neutralized. Similarly, xe2x80x9cnon-toxicxe2x80x9d with respect to the neutralizers means low or no toxicity to humans and other mammals, and does not mean non-toxic to the microorganisms being neutralized. xe2x80x9cSubstantial destructionxe2x80x9d of biological activity means at least as much destruction as is caused by porphyrin and porphyrin derivatives, metabolites and precursors which are known to have a damaging effect on biologically active proteins and cells of humans and mammals. Similarly, xe2x80x9csubstantially non-toxicxe2x80x9d means less toxic than porphyrin, porphyrin derivatives, metabolites and precursors that are known for blood sterilization. Preferably, neutralizers are less toxic than porphyrin, porphyrin derivatives, metabolites and precursors that are known for blood sterilization.
The term xe2x80x9cblood productxe2x80x9d as used herein includes blood constituents and therapeutic protein compositions containing proteins derived from blood as defined above. Fluids containing biologically active proteins other than those derived from blood may also be treated by the methods of this invention. Such fluids may also contain one or more components selected from the group consisting of protein, e.g. biologically active protein such as a therapeutic protein, blood and blood constituents, without destroying the biological activity of such components.
Decontamination methods of this invention using isoalloxazine derivatives as defined above do not substantially destroy the biological activity of fluid components other than microorganisms. As much biological activity of these components as possible is retained, although in certain instances, when the methods are optimized, some loss of biological activity, e.g., denaturization of protein components, must be balanced against effective decontamination of the fluid. So long as fluid components retain sufficient biological activity to be useful for their intended or natural purposes, their biological activities are not considered to be substantially destroyed.
xe2x80x9cDecompositionxe2x80x9d of the neutralizer upon exposure to light refers to the chemical transformation of the neutralizer into new compounds. An example of decomposition of the neutralizer is the production of lumichrome upon exposure of riboflavin to visible light
A xe2x80x9cphotosensitizerxe2x80x9d is defined as any compound which absorbs radiation of one or more defined wavelengths and subsequently utilizes the absorbed energy to carry out a chemical process. Photosensitizers of this invention may include compounds which preferentially adsorb to nucleic acids, thus focusing their photodynamic effect upon microorganisms and viruses with little or no effect upon accompanying cells or proteins. Other photosensitizers of this invention are also useful, such as those using singlet oxygen-dependent mechanisms.
An xe2x80x9calkylating agentxe2x80x9d is a compound that reacts with amino acid residues and nucleic bases and inhibits replication of microorganisms.